Comparative metabolism of 7,12-dimethylbenz[a]-anthracene and its non-carcinogenic 2-fluoro analogue by Syrian hamster embryo cells

The metabolism of 7,12-dimethylbenz[a]anthracene (DMBA) and its non-carcinogenic 2-fluoro analogue (2F-DMBA) by Syrian hamster embyro (SHE) cells has been studied using high pressure liquid chromatography (HPLC) and fluorescence spectroscopy. Metabolites produced by SHE cells were compared chromatographically to those produced on a larger scale by liver microsomal preparations and previously identified by gas chromatography and mass spectrometry. At least 2 (possibly 3) phenol metabolites, none of which appear to be in the A-ring, were formed from [3H] 2F-DMBA and totalled only 3% of the organic extractable activity present in the media at 24 h. On the other hand, 3 A-ring phenols (DMBA-2-ol, DMBA-3-ol and DMBA-4-ol) comprising almost 12% of the total organic extractable radioactivity at 24 h were identified as metabolites in SHE cell culture media. For both hydrocarbons the major organic extractable metabolite present at 24 h was the respective 8,9-dihydro-dihydroxydiol (DMBA 45%, 2F-DMBA 39%). Thus, substitution of fluorine for hydrogen at the 2-carbon of DMBA appears to block or greatly reduce the A-ring metabolism of this compound but has relatively little effect on D-ring oxidation. Therefore loss of the carcinogenic/mutagenic activity of DMBA correlates with the extent of A-ring metabolism including, possibly, the bay region diol epoxide.     

Effect of ara-A on differentiation and proliferation of HL-60 cells

The HL-60 human leukemic promyelocytic cell line can be induced to mature into terminally differentiated cells using certain chemotherapeutic agents. We have recently demonstrated that two inhibitors of DNA synthesis, cytosine arabinoside (ara-C) and aphidicolin, can induce HL-60 differentiation with the appearance of monocytic markers. These pyrimidine antimetabolites may have affected DNA methylation patterns and resulted in altered gene expression, or the differentiated phenotype may have occurred by inhibition of DNA replication. Consequently, we have extended these studies by using the purine analog, adenine arabinoside (ara-A), which also acts as an inhibitor of DNA synthesis. The results demonstrate that ara-A also induces HL-60 non-specific esterase activity and enhances expression of myeloid cell surface antigens, MY-4 and MO-1. The induction of a differentiated phenotype by ara-A occurs after partial inhibition of DNA synthesis, a finding similar to that observed with ara-C and aphidicolin. These observations indicate that purine, as well as pyrimidine analog inhibitors of DNA polymerization can induce differentiation of HL-60 cells along a monocytic lineage. These findings may be relevant to recent clinical trials that have employed low doses of ara-C in an attempt to induce differentiation of malignant hematopoietic cells.     

A simple method for exact intratracheal instillation in small laboratory rodents

An accurate, simple and quick method for instillation of substances into the respiratory tract of small rodents is reported. This new method is compared with conventional techniques.     

Identification and characterization of a human thymus-leukemia antigen (43-kDa/513TL) bearing a structural relationship to HLA

A human thymus-leukemia-like antigen has been identified that is antigenically distinct from T6/HTA-1. This was accomplished with a rabbit antiserum (513) which was prepared against lymphoblasts that were E rosette negative (E-), human thymus antigen positive (HuTA+), cALLA-, DR-, SmIg- from a patient who presented with a mediastinal mass and a WBC count of 130 X 10(9) cells/1. Following absorption with B cell and "null" cell lines, 513 exhibited prominent reactivity with a membrane antigen that was present on normal thymocytes and lymphoblasts from 11 of 13 patients with T cell ALL and 1 of 16 patients with common ALL, but was not detected on normal peripheral blood lymphocytes, normal bone marrow cells and leukemic lymphoblasts with an undifferentiated phenotype. SDS-PAGE analysis of this antigen indicated that it was composed of two subunits, 43-kDa and 12-kDa. Sequential absorption experiments revealed that: (1) the 12-kDa subunit was antigenically similar to beta 2 microglobulin; (2) the intact molecule did not exhibit HLA-A, B or C "framework" determinants; (3) the molecule was antigenically distinct from a human thymus-leukemia antigen HTA-1 (recognized by monoclonal antibodies NA1/34 and OKT6); and (4) the molecule was antigenically distinct from adenosine deaminase. It is concluded that 513 reacts with a membrane protein (designated 513TL) which exhibits properties characteristic of a histocompatibility-like antigen whose expression is restricted to thymocytes and some leukemias (TL antigen). Its antigenic distinction from another recently characterized human TL antigen, HTA-1, suggests polymorphism among this family of alloantigens.     

Cannabis Use,Lung Cancer,and Related Issues

The cannabis plant and its derivatives have been exploited for centuries for recreational and medicinal purposes, with millions of regular users around the world. The recreational use of cannabis is reflective of its neuropsychiatric effects, such as anxiolysis and euphoria. However, cannabis appears to have an emerging therapeutic role, especially in chronic disease and as an adjunct to cancer treatment. Increasing evidence supports cannabis in the management of chemotherapy-induced nausea and vomiting (CINV) and for pain management; however, studies are limited, particularly by difficulties associated with standardized dosing estimates and inability to accurately assess biologic activities of compounds in cannabis and derivative products. Smoking cannabis has not been proved to be a risk factor in the development of lung cancer, but the data are limited by small studies, misclassification due to self-reporting of use, small numbers of heavy cannabis smokers, and confounding of the risk associated with known causative agents for lung cancer (such as parallel chronic tobacco use). Cannabis and its biologically effective derivatives warrant additional research, ideally, controlled trials in which the cannabidiol and the delta-9-tetrahydrocabinol strength and use are controlled and documented.     

The CFU-S redistribution in spontaneous AKR leukemia

The number of bone marrow CFU-S decreases at the terminal stage of spontaneous AKR leukemia. This decrease is compensated by an increase of CFU-S in the spleen, lymph nodes, liver and thymus. Thus, the overall CFU-S number is basically equivalent to that found in normal mice. We take this as evidence of a spatial redistribution of CFU-S in AKR leukemia.     

Role of interleukin-2 in depressed T-cell mitogenesis after allogeneic marrow transplantation in man

Proliferative responses of mononuclear cells in liquid cultures to phytohemagglutinin, a T-cell mitogen, are depressed for a long time after allogeneic marrow transplantation. We examined the role of interleukin-2, a lymphokine important in T-cell mitogenesis, in the impaired phytohemagglutinin responses early after marrow transplantation. We found that interleukin-2 production, upon optimal stimulation, was impaired for mononuclear cells of most recipients early after marrow transplant. Further, we found that exogenous interleukin-2 did not restore depressed phytohemagglutinin responses of marrow transplant recipient mononuclear cells to normal. We hypothesize that early after allogeneic marrow transplant the helper T-cell pool is defective both in numbers, as shown by previous phenotypic studies, and in functional capabilities. There appear to be defects in both interleukin-2 production and interleukin-2 responsiveness in this system early after marrow transplant.     

Blastic transformation of a well-differentiated monocytic leukemia: Changes in cytochemical and cell surface markers

The clinical course of a patient with a well-differentiated monocytic leukemia which later underwent blastic transformation is described. Cytochemical, ultrastructural and cell surface analysis data were obtained at periods throughout her illness and correlated with the blastic transformation. Although surface markers characteristic of monocytic leukemia persisted, a deficiency of peroxidase in the granules of this patient's monocytes was observed as well as loss of α-naphthyl butyrate esterase staining during transformation.     

Nitrous oxide reduces growth of experimental rat leukemia

The ability of nitrous oxide to inhibit the in vivo growth of hematological neoplasms was investigated in a rat model for acute myeloid leukemia (BNML). Nitrous oxide, administered in a concentration of 67% with 33% oxygen, resulted in a reduction of spleen and liver weights of approx. 30%, as compared with leukemic rats kept in ambient air. Peripheral white cell counts were also considerably lower in the treated rats. Plasma levels of vitamin B₁₂ were found to be elevated in untreated leukemia, but fell to about normal levels after nitrous oxide exposure. On the contrary, folic acid levels were low in untreated leukemic rats, and significantly higher in animals exposed to nitrous oxide. The observed effects of nitrous oxide appeared to be dose-dependent. The deoxyuridine suppression test performed with leukemic cells became abnormal after nitrous oxide inhalation, in accordance with the effect on normal bone marrow. These results indicate that the interference of nitrous oxide with vitamin B₁₂-related metabolism, which leads to impairment of de novo thymidine synthesis, has the potency to reduce leukemic proliferation in vivo.     

Tumour initiation in mouse skin by 7,12-dimethylbenz[a]anthracene: irrelevance of systemic activation

The effect was investigated of 7,8-benzoflavone (BF) on tumour initiation by 7,12-dimethylbenz[a]anthracene (DMBA) applied either intragastrically or topically. A single topical application of BF drastically decreased tumour initiation via both routes. This decrease was in both cases inversely proportional to the interval between administration of DMBA and BF. The data suggest that the decisive step of metabolic activation of DMBA is mediated by cytochrome P-448 dependent monooxygenase(s) located within the epidermis.     

Extent of skin tumour initiation in mice by 7,12-dimethylbenz[a]anthracene and induction of arylhydrocarbon monooxygenase are not causally related

In order to clarify further the relation between extent of arylhydrocarbon monooxygenase (AHM) induction and tumour initiation by polycyclic aromatic hydrocarbons, AHM activity and concentration of 7,12-dimethylbenz[a]anthracene (DMBA) equivalents in dorsal epidermis were determined after both intragastric and topical administration of DMBA. Although in each case the dose applied caused comparable tumor incidences, both the extent of AHM induction and the concentration of DMBA equivalents were remarkably different: while topically applied DMBA stimulated AHM several-fold, intragastric instillation led to only a slight increase, if any. The concentration of DMBA equivalents in epidermis was about 100-fold higher after topical DMBA application than after intragastric administration.These data provide further evidence that extent of tumour initiation by DMBA and of AHM induction are not causally related; positive correlations found in certain cases appear to be fortuitous. Induction of AHM seems to be rather a function of DMBA concentration adjustment in epidermis and to indicate, if anything, elevated detoxification.     

A monoclonal antibody (SJ-9A4) to P24 present on common alls, neuroblastomas and platelets - II. Characterization of P24 and shedding in vitro and in vivo

The release of soluble P24 antigen into culture medium by common acute lymphoblastic leukemia (C-ALL) and neuroblastoma (NB) cell lines was studied. P24 release by C-ALL cells was detected using a solid phase indirect radioimmunometric assay (IRA) which combines the specificity of lectins and monoclonal antibodies (MoAb) and using immunoadsorption of labeled P24 in spent medium from cells incubated with 35S-methionine (met). No P24 was present in the medium of cells pulse labeled at 37 degrees C when they were placed at 4 degrees C, thus this is an active process. P24 release by NB cells could not be detected by IRA, but could be detected by immunoadsorption of spent medium of metabolically-labeled cells. The absence of IRA activity of P24 from NB spent medium was due to decreased glycosylation and thus no binding to the lectins employed in the IRA was observed. This was confirmed by lectin affinity chromatography which showed that P24 in the spent medium from C-ALL cells bound Ricinus communis agglutinin (RCA1), wheat germ agglutinin (WGA), concanavalin A (Con A), and lentil lectin (LcH), but not peanut agglutinin (PNA). P24 from NB cell spent medium did not bind to any of these lectins. The lectin affinity of P24 derived from lymphoblasts is consistent with the presence of N-linked oligosaccharide chains having N-acetyl glucosamine residues, a mannose core, and a terminal D-galactose. P24 from C-ALL cell spent medium was present in the 35-45% fraction of a saturated ammonium sulfate (SAS) partition of spent medium. The P24 antigen was detected in the fractionated plasma of five patients with C-ALL at the time of diagnosis and was undetectable when the patients had achieved a complete remission. Plasma from 2 patients with P24 negative ALL, normal human plasma, and normal human serum had no detectable activity.     

Adult T-cell leukemia: a report on two white patients

Two white European males are reported with adult T-cell leukemia (ATL), a disease first described in Japan, but recently also in the U.K. and U.S.A. Both patients presented with lymphadenopathy, but without a mediastinal mass. In addition, one patient had skin infiltrates and the other had hepatosplenomegaly. Morphologic and ultrastructural examination of the blasts in bone marrow and lymph node biopsy revealed a predominance of polymorphic lymphoid cells with pronounced nuclear irregularities and a semi-mature chromatine pattern. Histopathology of the lymph nodes showed a diffuse infiltration with medium-sized lymphoblasts with irregular nuclei. The blasts in the bone marrow formed E rosettes with sheep erythrocytes, lacked terminal deoxynucleotidyl transferase (Tdt) activity but expressed the Ia-like antigen; although the majority of the cells reacted with a polyclonal anti-T-cell serum, they were negative for OKT3. In one patient a helper/inducer phenotype (OKT4+) was found in the lymphoblasts of bone marrow and lymph node, while in the other only in the lymph node. The difference between bone marrow and lymph node phenotype is discussed. To our knowledge, these are the first two European patients reported with ATL, a disease clearly different from convoluted T-cell acute lymphocytic leukemia.     

A monoclonal antibody (SJ-9A4) to P24 present on common alls, neuroblastomas and platelets - I. Characterization and development of a unique radioimmunometric assay

We report the development and characterization of SJ-9A4, a monoclonal antibody (MoAb) produced against common acute lymphoblastic leukemia (C-ALL) cell lines. SJ-9A4 reacted with C-ALL, B-cell chronic lymphocytic leukemia (B-CLL), platelets and C-ALL neuroblastoma (NB) and the K562 cell lines. It had no significant reactivity with erythrocytes, granulocytes, circulating T or B lymphocytes, monocytes, granulocytic cell lines or a Ewing's sarcoma cell line. SJ-9A4 was shown to recognize the same region as two other MoAb to the p24 antigen, BA-2 and DU-ALL-1, as demonstrated by their ability to inhibit the binding of labeled SJ-9A4 to NALM-1 and NB cells. Other MoAb: J5, PI 153/3 and monoclonal anti-HLA-DR antibodies gave no inhibition. A solid phase indirect radioimmunometric assay (IRA) was developed which enabled the detection of P24 from C-ALL cells, utilizing its ability to bind the Ricinus communis agglutinin (RCA1) or wheat germ agglutinin (WGA) and SJ-9A4 simultaneously. When BA-2 and DU-ALL-1 were used in place of SJ-9A4, similar IRA results were obtained. Using the RCA1/SJ-9A4-IRA, P24 from as few as 1.6 X 10(4) cells of a C-ALL cell line could be detected; however, similar extracts of NB cell lines were negative despite high levels of SJ-9A4 binding to intact cells. The presence of P24 in NB extracts was demonstrated by (1) preincubation of NB extracts with SJ-9A4 which blocked MoAb binding to P24 and (2) immunoadsorption of P24 from solubilized membranes of 35S-methionine (met) labeled NB cells. Treatment of NB cells with neuraminidase did not result in IRA binding when either RCA1 or WGA were used as the solid phase lectin indicating that the differences in lectin affinity are not due to over sialation of NB membrane glycoproteins. These findings demonstrate a difference in the glycosylation of P24 from C-ALL and NB cells.     

The relationship of blast cell glucocorticoid receptor levels to response to single-agent steroid trial and remission response in children with acute lymphoblastic leukemia

Of 263 children with glucocorticoid receptor (GR) levels measured at diagnosis of acute lymphoblastic leukemia (ALL), 27 received single-agent glucocorticoid before combination induction chemotherapy and were evaluable for in vivo clinical response to steroid. Twenty-one were glucocorticoid-responsive and 6 were resistant. There was no difference between the two groups in the distribution of age, sex, white blood cell count, immunophenotype of blasts, initial central nervous system disease or mediastinal mass. The median GR level, however, was appreciably lower in the group of patients with resistant disease (6250 vs 17,800 sites/cell, p = 0.06). Five of 12 patients with GR levels of less than 10,000 sites/cell compared to 1 of 15 with higher levels had glucocorticoid-resistant ALL (p = 0.03). All 21 patients with glucocorticoid-sensitive disease achieved a complete remission after combination induction chemotherapy, but only 3 of 5 evaluable patients in the other group did (p less than 0.04). Two patients were studied both at diagnosis and at relapse; both had decreased GR levels at relapse (below detection in one) and failed to respond to glucocorticoid. We conclude that a lower GR level is associated with glucocorticoid resistance and furthermore that a decrease in the level of GR is a mechanism of acquiring steroid resistance.     

Thymic stroma in AKR mice: its function and virus production

This study reports experiments with thymic stromal remnants in AKR mice, a strain with a high natural incidence of thymic lymphoma. A method has been developed in which thymic stromal cells which survive a 4-week culture period, 1 week at 24 degrees C and 3 weeks at 37 degrees C are suitable for grafting. Most thymic lymphocytes die under these conditions. Stromal remnants were studied by culturing and grafting under the kidney capsule of 2-month-old syngeneic mice. Their in vitro morphology and virus production, their ability to reconstitute a new thymus from host progenitors and their eventual lymphoma development was evaluated. The stromal remnants were from: 1- and 3-month-old normal mice; 6-10-month-old normal mice; 21-28-day-old animals treated with the lymphomagenic virus, SL3-3, at 3 days of age. Our data show that thymic stromal function as measured by lymphoid reconstitution of thymic stromal grafts of AKR mice is not impaired with age or by the presence of oncogenic virus. Oncogenic viruses are found in the thymic stroma of old mice and in thymic stroma of young virus-treated mice. Oncogenic viruses are not found in thymic stroma of young normal mice. Lymphoma can develop in the grafted stromal remnants expressing lymphomagenic virus.     

Bone marrow transplantation performed as first-time treatment in two cases of secondary acute myeloid leukemia

Two young patients with secondary acute myeloid leukemia were treated with allogeneic bone marrow transplantation as first-line treatment for their disease. High dose cytosine arabinoside was added to the conventional preparative regimen of cyclophosphamide and total body irradiation. No major problems were encountered to the immediate post-transplantation period. Complete remission of the acute myeloid leukemia was noted in both patients, with no evidence of recurrent disease at 15+ and 13 months. One patient developed severe pancytopenia 10 months after grafting with the presence of antibodies against platelets and granulocytes from the bone marrow donor. She died 3 months later from a generalised Aspergillus septicemia, probably precipitated by therapy with high doses of methylprednisolone. The other patient is alive in excellent clinical condition and in hematologic remission. Bone marrow transplantation must be taken into consideration as first-line therapy in any young patient who is suffering from a refractory type of leukemia and who has a suitable donor.